In Europe, sand flies (Diptera: Psychodidae: Phlebotominae) are characteristic Mediterranean fauna, though some species expand their range further north. However, the sand fly fauna of Central Europe remains underreported, particularly in Hungary where recent data is lacking due to limited and outdated entomological surveys. To address this gap, a series of sand fly surveys were conducted in Hungary, with significant findings from two trapping efforts in 2017 and 2024. In 2017, only a single female Phlebotomus papatasi was trapped in northern Hungary, which marks one of the northernmost records of the species. In 2024, a more extensive and geographically wider survey recorded 264 sand flies at 34 sites, including three species: Ph. mascittii, Ph. neglectus, and Ph. papatasi. Sand flies were found across diverse environmental settings, including urban, agricultural, and natural habitats. Particularly, the previously rare presence of Ph. mascittii at rural sites (natural rock formations) was reported. Analysis of historical and current data revealed the presence of four sand fly species in Central and South Transdanubia, with evidence suggesting potential range expansion. Blood meal analysis of engorged females identified a variety of domestic and wild host species, but no Leishmania or Phlebovirus infections were detected. Habitat modelling and linear discriminant analysis indicated substantial climate suitability across Southeast Europe, with most positive sand fly observations observed in discontinuous urban fabric CORINE Land Cover classes. This study offers important insights into the ecology, distribution, and climatic preferences of sand flies in Hungary and provides crucial baseline data to monitor potential future spread.

• Keywords
• Barcoding, Blood meal, Central Europe, Ecology, MALDI-MS, Sand flies,
• Publication type
• Journal Article MeSH

BACKGROUND: Phlebotomine sand flies (Diptera: Psychodiae) in the Republic of Moldova have been understudied for decades. Our study provides a first update on their occurrence, species composition and bloodmeal sources after 50 years. METHODS: During 5 seasons (2013-2017), 58 localities from 20 regions were surveyed for presence of sand flies using CDC light traps and manual aspirators. Species identification was done by a combination of morphological and molecular approaches (DNA barcoding, MALDI-TOF MS protein profiling). In engorged females, host blood was identified by three molecular techniques (RFLP, cytb sequencing and MALDI-TOF peptide mass mapping). Population structure of most abundant species was studied by cox1 haplotyping; phylogenetic analyses of ITS2 and cox1 genetic markers were used to resolve relationships of other detected species. RESULTS: In total, 793 sand flies were collected at 30 (51.7%) localities from 12 regions of Moldova. Three species were identified by an integrative morphological and molecular approach: Phlebotomus papatasi, P. perfiliewi and Phlebotomus sp. (Adlerius), the first being the most abundant and widespread, markedly anthropophilic based on bloodmeal analyses, occurring also indoors and showing low population structure with only five haplotypes of cox1 detected. Distinct morphological and molecular characters of Phlebotomus sp. (Adlerius) specimens suggest the presence of a yet undescribed species. CONCLUSIONS: Our study revealed the presence of stable sand fly populations of three species in Moldova that represent a biting nuisance as well as a potential threat of pathogen transmission and shall be further studied.

• Keywords
• Bloodmeal analysis, Haplotype network, MALDI-TOF MS protein profiling, Phlebotomus,
• MeSH
• Phylogeny MeSH
• Haplotypes MeSH
• Insect Vectors classification   genetics   parasitology   MeSH
• Host Specificity MeSH
• Leishmaniasis transmission   MeSH
• Humans MeSH
• Phlebotomus classification   genetics   physiology   MeSH
• Psychodidae classification   genetics   physiology   MeSH
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Moldova epidemiology   MeSH

BACKGROUND: Ethiopia is affected by human leishmaniasis caused by several Leishmania species and transmitted by a variety of sand fly vectors of the genus Phlebotomus. The sand fly fauna in Ethiopia is highly diverse and some species are closely related and similar in morphology, resulting in difficulties with species identification that requires deployment of molecular techniques. DNA barcoding entails high costs, requires time and lacks reference sequences for many Ethiopian species. Yet, proper species identification is pivotal for epidemiological surveillance as species differ in their actual involvement in transmission cycles. Recently, protein profiling using MALDI-TOF mass spectrometry has been introduced as a promising technique for sand fly identification. METHODS: In our study, we used an integrative taxonomic approach to identify most of the important sand fly vectors of leishmaniasis in Ethiopia, applying three complementary methods: morphological assessment, sequencing analysis of two genetic markers, and MALDI-TOF MS protein profiling. RESULTS: Although morphological assessment resulted in some inconclusive identifications, both DNA- and protein-based techniques performed well, providing a similar hierarchical clustering pattern for the analyzed species. Both methods generated species-specific sequences or protein patterns for all species except for Phlebotomus pedifer and P. longipes, the two presumed vectors of Leishmania aethiopica, suggesting that they may represent a single species, P. longipes Parrot & Martin. All three approaches also revealed that the collected specimens of Adlerius sp. differ from P. (Adlerius) arabicus, the only species of Adlerius currently reported in Ethiopia, and molecular comparisons indicate that it may represent a yet undescribed new species. CONCLUSIONS: Our study uses three complementary taxonomical methods for species identification of taxonomically challenging and yet medically import Ethiopian sand flies. The generated MALDI-TOF MS protein profiles resulted in unambiguous identifications, hence showing suitability of this technique for sand fly species identification. Furthermore, our results contribute to the still inadequate knowledge of the sand fly fauna of Ethiopia, a country severely burdened with human leishmaniasis.

• Keywords
• DNA barcoding, Ethiopia, MALDI-TOF mass spectrometry, Morphology, Phlebotomus, Protein profiling, Sand flies,
• MeSH
• Species Specificity MeSH
• Phylogeny MeSH
• Insect Vectors classification   MeSH
• Leishmaniasis MeSH
• Psychodidae classification   MeSH
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Ethiopia epidemiology   MeSH

BACKGROUND: Sand flies (Diptera: Psychodidae) are medically important vectors of human and veterinary disease-causing agents. Among these, the genus Leishmania (Kinetoplastida: Trypanosomatidae), and phleboviruses are of utmost importance. Despite such significance, updated information about sand fly fauna is missing for Balkan countries where both sand flies and autochtonous leishmaniases are historically present and recently re-emerging. Therefore, a review of historical data on sand fly species composition and distribution in the region was followed by a large-scale entomological survey in eight Balkan countries to provide a recent update on local sand fly fauna. METHODS: The literature search involved the period 1910-2019. The entomological survey was conducted at 1189 sampling stations in eight countries (Bulgaria, Bosnia and Herzegovina, Croatia, Kosovo, Montenegro, North Macedonia, Serbia and Slovenia), covering 49 settlements and 358 sampling sites between June and October in the years 2014 and 2016, accumulating 130 sampling days. We performed a total of 1189 trapping nights at these stations using two types of traps (light and CO2 attraction traps) in each location. Sampling was performed with a minimal duration of 6 (Montenegro) and a maximal of 47 days (Serbia) between 0-1000 m.a.s.l. Collected sand flies were morphologically identified. RESULTS: In total, 8490 sand fly specimens were collected. Morphological identification showed presence of 14 species belonging to genera Phlebotomus and Sergentomyia. Historical data were critically reviewed and updated with our recent findings. Six species were identified in Bosnia and Herzegovina (2 new records), 5 in Montenegro (2 new records), 5 in Croatia (2 new records), 9 in Bulgaria (5 new records), 11 in North Macedonia (1 new record), 10 in Serbia (no new records), 9 in Kosovo (3 new records) and 4 in Slovenia (no new records). CONCLUSIONS: This study presents results of the first integrated sand fly fauna survey of such scale for the Balkan region, providing first data on sand fly populations for four countries in the study area and presenting new species records for six countries and updated species lists for all surveyed countries. Our findings demonstrate presence of proven and suspected vectors of several Leishmania species.

• Keywords
• Balkans, Morphology, Phlebotomine, Sand fly, Species,
• MeSH
• Insect Vectors classification   parasitology   MeSH
• Leishmaniasis transmission   MeSH
• Psychodidae classification   parasitology   MeSH
• Animal Distribution * MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Review MeSH
• Geographicals
• Balkan Peninsula MeSH